There has emerged considerable clinical and basic biological interest in the pathway of mitochondrial fatty acid B-oxidation, largely as a result of studies in patients with inherited defects in this pathway. Dr. Roe and his colleagues at Duke University and Dr. Coates at Children's Hospital of Philadelphia have identified a large cohort of children whose clinical presentations include hypotonia and developmental delay, cardiomyopathy, Reye- like illness, or sudden infant death. Both in vivo and in vitro biochemical data from these cases suggest impaired B-oxidation of saturated and unsaturated fatty acids and the likelihood of enzyme defects, as yet undefined, that would not be detected by current tests. Stable isotope-labelled unsaturated fatty acid precursors, made available by Dr. Sprecher at Ohio State University, will be used to probe the pathway of fatty acid oxidation in vitro using fibroblasts and in vivo in patients with a strong clinical suspicion of an abnormality in this pathway. The accumulated intermediates will precisely indicate the specific metabolite block in cells from patients with defects in fatty acid oxidation. Fibroblasts will be incubated with isotopically labeled 4-cis- decenoic acid because its complete oxidation requires the concerned action of all B-oxidation enzymes, including those required for unsaturated fatty acid oxidation, with the exception of long-chain acyl-CoA dehydrogenase. Samples of the incubation medium will be sent to Dr. Millington at Duke University for the gas chromatographic-mass spectrometric (GC-MS) analysis of fatty acid intermediates. Clinical studies of patients will use deuterium- labelled linoleic acid, a long-chain polyunsaturated fatty acid, to probe the pathway of B-oxidation in vivo. GC-MS and fast atom bombardment-MS techniques will be used to analyze plasma and urinary fatty acid intermediates and acylcarnitines. Since these disorders will share an inability to produce ketone bodies normally, treatment strategies involving restoration of this fuel source will be attempted with RS-1, 3-butanediol as an alternate dietary source. The major objectives, therefore, are to characterize and demonstrate new defects of fat oxidation and to investigate a potential therapy.